Abstract
Reverse transcription polymerase chain reaction (RT-PCR) for BCR::ABL1 is the most common and widely accepted method of measurable residual disease (MRD) assessment in Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL); however, RT-PCR may not be an optimal measure of MRD in many cases of Ph+ ALL. We evaluated the clinical impact of a highly sensitive next-generation sequencing (NGS) MRD assay (sensitivity of 10−6) and its correlation with RT-PCR for BCR::ABL1 in patients with Ph+ ALL. Overall, 32% of patients had a discordance between MRD assessment by RT-PCR and NGS, and 31% of patients who achieved NGS MRD negativity were PCR+ at the same timepoint. Among eight patients with long-term detectable BCR::ABL1 by PCR, six were PCR+/NGS−. These patients generally had stable PCR levels that persisted despite therapeutic interventions, and none subsequently relapsed; in contrast, patients who were PCR+/NGS+ had more variable PCR values that responded to therapeutic intervention. In a separate cohort of prospectively collected clinical samples, 11 of 65 patients (17%) with Ph+ ALL who achieved NGS MRD negativity had detectable BCR::ABL1 by PCR, and none of these patients relapsed. Relapse-free survival and overall survival were similar in patients who were PCR+/NGS− and PCR−/NGS−, suggesting that PCR for BCR::ABL1 did not provide additional prognostic information in patients who achieved NGS MRD negativity. NGS-based assessment of MRD is prognostic in Ph+ ALL and identifies patients with low-level detectable BCR::ABL1 who are unlikely to relapse nor to benefit from therapeutic interventions.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1196-1203 |
| Number of pages | 8 |
| Journal | American journal of hematology |
| Volume | 98 |
| Issue number | 8 |
| DOIs | |
| State | Published - Aug 2023 |
ASJC Scopus subject areas
- Hematology
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In: American journal of hematology, Vol. 98, No. 8, 08.2023, p. 1196-1203.
Research output: Contribution to journal › Article › peer-review
}
TY - JOUR
T1 - Ultrasensitive NGS MRD assessment in Ph+ ALL: Prognostic impact and correlation with RT-PCR for BCR::ABL1
AU - Short, Nicholas J.
AU - Jabbour, Elias
AU - Macaron, Walid
AU - Ravandi, Farhad
AU - Jain, Nitin
AU - Kanagal-Shamanna, Rashmi
AU - Patel, Keyur P.
AU - Loghavi, Sanam
AU - Haddad, Fadi G.
AU - Yilmaz, Musa
AU - Issa, Ghayas C.
AU - Kebriaei, Partow
AU - Kornblau, Steven M.
AU - Pelletier, Sarah
AU - Flores, Wilmer
AU - Matthews, Jairo
AU - Garris, Rebecca
AU - Kantarjian, Hagop
N1 - Funding Information: N.J.S has received honoraria from Adaptive Biotechnologies Co. E.J. has received research funding and has received honoraria from Adaptive Biotechnologies Co. The other authors report no relevant conflicts of interest. Adaptive Biotechnologies Co. performed the NGS MRD assay at no cost to the authors. Supported by an MD Anderson Cancer Center Support Grant (CA016672) and SPORE. N.J.S. is supported by the American Society of Hematology Junior Faculty Scholar Award in Clinical Research. The clonoSEQ MRD assay (Adaptive Biotechnologies Co., Seattle, WA) is an NGS-based immunosequencing assay that was used for this analysis, as previously described.9 Briefly, pretreatment genomic DNA derived from stored bone marrow specimens was sequenced using multiplex PCR assays composed of primers targeted to the variable (V) genes (forward primers) and joining (J) genes (reverse primers) of the CDR3 region within IG and TR genes. Samples for which a trackable B-cell receptor sequence was not identified were reflexed for analysis of TRG followed subsequently by TRB. Using high-throughput NGS, the index sequence(s) identified in the diagnostic sample were specifically searched for and quantified, if present, in the remission bone marrow. This assay has an analytic sensitivity of 0.0001% (1 × 10−6), with a sensitivity that is dependent on the cellular input. NGS-based MRD assessment was performed retrospectively for the primary analyses, and this information was therefore not available to clinicians for decision-making. Analysis of a separate validation cohort of prospectively collected clinical NGS MRD assessments was also performed (see section below: “Validation of discordance between RT-PCR for BCR::ABL1 and NGS MRD in prospectively collected clinical samples”), and this information was available in real time to the treating clinician. These latter samples may have been obtained from either the bone marrow or peripheral blood. For all analyses comparing PCR and NGS MRD assessments, samples for both assays were collected at the same timepoint. A flowchart of the study and patient samples in the two cohorts is shown in Figure S1. This is a retrospective study evaluating the prognostic impact of a highly sensitivity NGS MRD assay in patients with Ph+ ALL. Eligible patients received frontline ALL therapy at our institution between March 2006 and June 2019 using a backbone of hyper-CVAD-based therapy plus a BCR::ABL1 TKI and achieved complete remission as best response. Patients who received frontline blinatumomab-based therapies were excluded. In the primary analysis, patients were selected for inclusion based on the availability of a banked pretreatment bone marrow sample (for purposes of clonality determination) and at least 1 post-treatment remission bone marrow sample that met one or more of three prespecified criteria: (1) remission marrow collected between 1 month and 6 months from the start of frontline treatment, regardless of MRD response by RT-PCR for BCR::ABL1 (“early MRD” group), (2) remission marrow collected >2 years from the start of treatment in a patient who had been in CMR for at least 1 year (“long-term PCR−” group), and (3) remission marrow collected >2 years from the start of treatment in a patient who had persistently detectable, stable PCR values for at least 1 year (“long-term PCR+” group). For criteria 2–3, patients were required to be in continuous first remission without prior alloSCT. This study was conducted at a single academic center (The University of Texas MD Anderson Cancer Center [UTMDACC]). This study was approved by the Institutional Review Board of UTMDACC and was conducted in accordance with the Declaration of Helsinki. MRD assessment was performed in our clinical laboratory on fresh bone marrow samples or peripheral blood, as previously described.17 The sensitivity of this assay is between 0.01% (10−4) and 0.001% (10−5). Per our institutional practice, RT-PCR for BCR::ABL1 for MRD was assessed with each remission bone marrow, approximately every 1–3 months for the first year of treatment, then approximately every 3–6 months thereafter. RT-PCR MRD information was available from the clinical laboratory and was available to clinicians for decision-making. The clonoSEQ MRD assay (Adaptive Biotechnologies Co., Seattle, WA) is an NGS-based immunosequencing assay that was used for this analysis, as previously described.9 Briefly, pretreatment genomic DNA derived from stored bone marrow specimens was sequenced using multiplex PCR assays composed of primers targeted to the variable (V) genes (forward primers) and joining (J) genes (reverse primers) of the CDR3 region within IG and TR genes. Samples for which a trackable B-cell receptor sequence was not identified were reflexed for analysis of TRG followed subsequently by TRB. Using high-throughput NGS, the index sequence(s) identified in the diagnostic sample were specifically searched for and quantified, if present, in the remission bone marrow. This assay has an analytic sensitivity of 0.0001% (1 × 10−6), with a sensitivity that is dependent on the cellular input. NGS-based MRD assessment was performed retrospectively for the primary analyses, and this information was therefore not available to clinicians for decision-making. Analysis of a separate validation cohort of prospectively collected clinical NGS MRD assessments was also performed (see section below: “Validation of discordance between RT-PCR for BCR::ABL1 and NGS MRD in prospectively collected clinical samples”), and this information was available in real time to the treating clinician. These latter samples may have been obtained from either the bone marrow or peripheral blood. For all analyses comparing PCR and NGS MRD assessments, samples for both assays were collected at the same timepoint. A flowchart of the study and patient samples in the two cohorts is shown in Figure S1. CMR was defined as the absence of a quantifiable BCR::ABL1 transcript by RT-PCR, as previously described.3 Relapse was defined as recurrence of bone marrow blasts >5% or extramedullary ALL. Relapse-free survival (RFS) was calculated from the time of response until relapse or death from any cause, censored if the patient was alive at last follow-up. Overall survival (OS) was calculated from the time of treatment initiation until death from any cause, censored if the patient was alive at last follow-up. Survival estimates were not censored at the time of alloSCT. Patient characteristics were summarized using median (range) for continuous variables and frequencies (percentages) for categorical variables. To compare two groups with continuous variables, the Wilcoxon rank-sum test was performed. The Kaplan–Meier method was used to estimate the probabilities for RFS and OS and differences between groups were evaluated with the log-rank test. All statistical analyses were performed using GraphPad Prism 9. This is a retrospective study evaluating the prognostic impact of a highly sensitivity NGS MRD assay in patients with Ph+ ALL. Eligible patients received frontline ALL therapy at our institution between March 2006 and June 2019 using a backbone of hyper-CVAD-based therapy plus a BCR::ABL1 TKI and achieved complete remission as best response. Patients who received frontline blinatumomab-based therapies were excluded. In the primary analysis, patients were selected for inclusion based on the availability of a banked pretreatment bone marrow sample (for purposes of clonality determination) and at least 1 post-treatment remission bone marrow sample that met one or more of three prespecified criteria: (1) remission marrow collected between 1 month and 6 months from the start of frontline treatment, regardless of MRD response by RT-PCR for BCR::ABL1 (“early MRD” group), (2) remission marrow collected >2 years from the start of treatment in a patient who had been in CMR for at least 1 year (“long-term PCR−” group), and (3) remission marrow collected >2 years from the start of treatment in a patient who had persistently detectable, stable PCR values for at least 1 year (“long-term PCR+” group). For criteria 2–3, patients were required to be in continuous first remission without prior alloSCT. This study was conducted at a single academic center (The University of Texas MD Anderson Cancer Center [UTMDACC]). This study was approved by the Institutional Review Board of UTMDACC and was conducted in accordance with the Declaration of Helsinki. Funding Information: Adaptive Biotechnologies Co. performed the NGS MRD assay at no cost to the authors. Supported by an MD Anderson Cancer Center Support Grant (CA016672) and SPORE. N.J.S. is supported by the American Society of Hematology Junior Faculty Scholar Award in Clinical Research. Publisher Copyright: © 2023 Wiley Periodicals LLC.
PY - 2023/8
Y1 - 2023/8
N2 - Reverse transcription polymerase chain reaction (RT-PCR) for BCR::ABL1 is the most common and widely accepted method of measurable residual disease (MRD) assessment in Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL); however, RT-PCR may not be an optimal measure of MRD in many cases of Ph+ ALL. We evaluated the clinical impact of a highly sensitive next-generation sequencing (NGS) MRD assay (sensitivity of 10−6) and its correlation with RT-PCR for BCR::ABL1 in patients with Ph+ ALL. Overall, 32% of patients had a discordance between MRD assessment by RT-PCR and NGS, and 31% of patients who achieved NGS MRD negativity were PCR+ at the same timepoint. Among eight patients with long-term detectable BCR::ABL1 by PCR, six were PCR+/NGS−. These patients generally had stable PCR levels that persisted despite therapeutic interventions, and none subsequently relapsed; in contrast, patients who were PCR+/NGS+ had more variable PCR values that responded to therapeutic intervention. In a separate cohort of prospectively collected clinical samples, 11 of 65 patients (17%) with Ph+ ALL who achieved NGS MRD negativity had detectable BCR::ABL1 by PCR, and none of these patients relapsed. Relapse-free survival and overall survival were similar in patients who were PCR+/NGS− and PCR−/NGS−, suggesting that PCR for BCR::ABL1 did not provide additional prognostic information in patients who achieved NGS MRD negativity. NGS-based assessment of MRD is prognostic in Ph+ ALL and identifies patients with low-level detectable BCR::ABL1 who are unlikely to relapse nor to benefit from therapeutic interventions.
AB - Reverse transcription polymerase chain reaction (RT-PCR) for BCR::ABL1 is the most common and widely accepted method of measurable residual disease (MRD) assessment in Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL); however, RT-PCR may not be an optimal measure of MRD in many cases of Ph+ ALL. We evaluated the clinical impact of a highly sensitive next-generation sequencing (NGS) MRD assay (sensitivity of 10−6) and its correlation with RT-PCR for BCR::ABL1 in patients with Ph+ ALL. Overall, 32% of patients had a discordance between MRD assessment by RT-PCR and NGS, and 31% of patients who achieved NGS MRD negativity were PCR+ at the same timepoint. Among eight patients with long-term detectable BCR::ABL1 by PCR, six were PCR+/NGS−. These patients generally had stable PCR levels that persisted despite therapeutic interventions, and none subsequently relapsed; in contrast, patients who were PCR+/NGS+ had more variable PCR values that responded to therapeutic intervention. In a separate cohort of prospectively collected clinical samples, 11 of 65 patients (17%) with Ph+ ALL who achieved NGS MRD negativity had detectable BCR::ABL1 by PCR, and none of these patients relapsed. Relapse-free survival and overall survival were similar in patients who were PCR+/NGS− and PCR−/NGS−, suggesting that PCR for BCR::ABL1 did not provide additional prognostic information in patients who achieved NGS MRD negativity. NGS-based assessment of MRD is prognostic in Ph+ ALL and identifies patients with low-level detectable BCR::ABL1 who are unlikely to relapse nor to benefit from therapeutic interventions.
UR - http://www.scopus.com/inward/record.url?scp=85159195958&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85159195958&partnerID=8YFLogxK
U2 - 10.1002/ajh.26949
DO - 10.1002/ajh.26949
M3 - Article
C2 - 37183966
AN - SCOPUS:85159195958
SN - 0361-8609
VL - 98
SP - 1196
EP - 1203
JO - American journal of hematology
JF - American journal of hematology
IS - 8
ER -